			Home About us Contact

Metabolic Profiling (metabolic + profiling)

Distribution by Scientific Domains

Life Sciences	52%
Chemistry	38%
Medical Sciences	9%

Selected Abstracts

Stimulation, Monitoring, and Analysis of Pathway Dynamics by Metabolic Profiling in the Aromatic Amino Acid Pathway

BIOTECHNOLOGY PROGRESS, Issue 6 2004
M. Oldiges
Using a concerted approach of biochemical standard preparation, analytical access via LC-MS/MS, glucose pulse, metabolic profiling, and statistical data analysis, the metabolism dynamics in the aromatic amino acid pathway has been stimulated, monitored, and analyzed in different tyrosine-auxotrophic l -phenylalanine-producing Escherichiacoli strains. During the observation window from ,4 s (before) up to 27 s after the glucose pulse, the dynamics of the first five enzymatic reactions in the aromatic amino acid pathway was observed by measuring intracellular concentrations of 3-deoxy- d -arabino-heptulosonate 7-phosphate DAH(P), 3-dehydroquinate (3-DHQ), 3-dehydroshikimate (3-DHS), shikimate 3-phosphate (S3P), and shikimate (SHI), together with the pathway precursors phosphoenolpyruvate (PEP) and P5P, the lumped pentose phosphate pool as an alternative to the nondetectable erythrose 4-phosphate (E4P). Provided that a sufficient fortification of the carbon flux into the pathway of interest is ensured, respective metabolism dynamics can be observed. On the basis of the intracellular pool measurements, the standardized pool velocities were calculated, and a simple, data-driven criterion-called "pool efflux capacity" (PEC)-is derived. Despite its simplifying system description, the criterion managed to identify the well-known AroB limitation in the E. coli strain A (genotype ,( pheA tyrA aroF)/pJF119EH aroFfbrpheAfbramp) and it also succeeded to identify AroL and AroA (in strain B, genotype ,( pheA tyrA aroF)/pJF119EH aroFfbrpheAfbraroB amp) as promising metabolic engineering targets to alleviate respective flux control in subsequent l -Phe producing strains. Furthermore, using of a simple correlation analysis, the reconstruction of the metabolite sequence of the observed pathway was enabled. The results underline the necessity to extend the focus of glucose pulse experiments by studying not only the central metabolism but also anabolic pathways. [source]

Effects of cevoglitazar, a dual PPAR,/, agonist, on ectopic fat deposition in fatty Zucker rats

DIABETES OBESITY & METABOLISM, Issue 6 2009
D. Laurent
Aim:, By acting as both insulin sensitizers and lipid-lowering agents, dual-acting peroxisome proliferator-activated receptors ,/, (PPAR,/,) agonists may be used to improve glucose tolerance in type 2 diabetic patients without inducing adiposity and body weight gain. Here, in an animal model of obesity and insulin resistance, the metabolic response to cevoglitazar, a dual PPAR,/,, was characterized using a combination of in vivo and ex vivo magnetic resonance methodologies and compared to treatment effects of fenofibrate, a PPAR, agonist, and pioglitazone, a PPAR, agonist. Methods:, Four groups of fatty Zucker rats: (i) Vehicle; (ii) fenofibrate 150 mg/kg; (iii) pioglitazone 30 mg/kg; and (iv) cevoglitazar 5 mg/kg were investigated before and after treatment. Animals were fed a fat-enriched (54% kcal fat) diet for 6 weeks, 2 weeks high of fat,exposure alone followed by a 4-week dosing period. Results and conclusions:, Cevoglitazar was as effective as pioglitazone at improving glucose tolerance. However, unlike pioglitazone, both fenofibrate and cevoglitazar reduced BW gain and adiposity, independent of food intake. All three treatment regimens normalized intramyocellular lipids. Metabolic profiling showed that in the muscle cevoglitazar improves the lipid profile via both PPAR,- and PPAR,-mediated mechanisms. Pioglitazone reduced hepatic lipid accumulation, while cevoglitazar and fenofibrate reduced hepatic lipid concentration below baseline levels (p < 0.05). Metabolic profiling showed that in the liver, cevoglitazar functions largely through PPAR, agonism resulting in increased ,-oxidation. Cevoglitazar only induced small changes to the lipid composition of visceral fat. In subcutaneous fat, however, cevoglitazar induced changes similar to those observed with fenofibrate suggesting export of fatty acids from this depot. [source]

Red Pyrroloquinoline Alkaloids from the Mushroom Mycena haematopus

EUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 2 2008
Silke Peters
Abstract Four so far unknown red alkaloid pigments, haematopodin B (1) and the mycenarubins D (3), E (5) and F (6), were isolated from fruiting bodies of Mycena haematopus. The structures of these pyrroloquinoline alkaloids were established by 2D NMR spectroscopic and MS (ESI) methods. Their absolute configurations were determined by comparison of their CD spectra with that of haematopodin (2) or mycenarubin A (4). Metabolic profiling of the red pigments of intact and injured fruiting bodies revealed that the known degradation product haematopodin (2) originates from haematopodin B (1), which is the native main pigment of M. haematopus.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008) [source]

TICL , a web tool for network-based interpretation of compound lists inferred by high-throughput metabolomics

FEBS JOURNAL, Issue 7 2009
Alexey V. Antonov
High-throughput metabolomics is a dynamically developing technology that enables the mass separation of complex mixtures at very high resolution. Metabolic profiling has begun to be widely used in clinical research to study the molecular mechanisms of complex cell disorders. Similar to transcriptomics, which is capable of detecting genes at differential states, metabolomics is able to deliver a list of compounds differentially present between explored cell physiological conditions. The bioinformatics challenge lies in a statistically valid interpretation of the functional context for identified sets of metabolites. Here, we present TICL, a web tool for the automatic interpretation of lists of compounds. The major advance of TICL is that it not only provides a model of possible compound transformations related to the input list, but also implements a robust statistical framework to estimate the significance of the inferred model. The TICL web tool is freely accessible at http://mips.helmholtz-muenchen.de/proj/cmp. [source]

Metabolic profiling as a tool for revealing Saccharomyces interactions during wine fermentation

FEMS YEAST RESEARCH, Issue 1 2006
Kate S. Howell
Abstract The multi-yeast strain composition of wine fermentations has been well established. However, the effect of multiple strains of Saccharomyces spp. on wine flavour is unknown. Here, we demonstrate that multiple strains of Saccharomyces grown together in grape juice can affect the profile of aroma compounds that accumulate during fermentation. A metabolic footprint of each yeast in monoculture, mixed cultures or blended wines was derived by gas chromatography , mass spectrometry measurement of volatiles accumulated during fermentation. The resultant ion spectrograms were transformed and compared by principal-component analysis. The principal-component analysis showed that the profiles of compounds present in wines made by mixed-culture fermentation were different from those where yeasts were grown in monoculture fermentation, and these differences could not be produced by blending wines. Blending of monoculture wines to mimic the population composition of mixed-culture wines showed that yeast metabolic interactions could account for these differences. Additionally, the yeast strain contribution of volatiles to a mixed fermentation cannot be predicted by the population of that yeast. This study provides a novel way to measure the population status of wine fermentations by metabolic footprinting. [source]

Development and validation of a gas chromatography/mass spectrometry metabonomic platform for the global profiling of urinary metabolites

RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 19 2008
Kishore K. Pasikanti
This paper presents a simple and reliable gas chromatography/mass spectrometry (GC/MS) method for the metabonomic analysis of human urine samples. The sample preparation involved the depletion of excess urea via treatment with urease and subsequent protein precipitation using ice-cold ethanol. An aliquot of the mixture was separated, dried, trimethylsilyl (TMS)-derivatized and 1.0,µL of the derivatized extract was injected into the GC/MS system via split injection (1:10). Approximately 150 putative metabolites belonging to different chemical classes were identified from the pooled human urine samples. All the identified metabolites were selected to evaluate precision and stability of the GC/MS assay. More than 95% of the metabolites demonstrated good reproducibility, with intra-day and inter-day precision values below 15%. Metabolic profiling of 53 healthy male and female urine samples in combination with pattern recognition techniques was performed to further validate the GC/MS metabolite profiling assay. Principal component analysis (PCA) followed by orthogonal partial least squares analysis (OPLS) revealed differences between urinary metabolite profiles of healthy male and female subjects. This validated GC/MS metabolic profiling method may be further applied to the metabonomic screening of urinary biomarkers in clinical studies. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Metabolic profiling as a tool for understanding defense response of Taxus Cuspidata cells to shear stress

BIOTECHNOLOGY PROGRESS, Issue 5 2009
Pei-Pei Han
Abstract To obtain a better understanding of responsive mechanism of plant cells in response to hydrodynamic mechanical stress, a metabolic profiling approach was used to profile metabolite changes of Taxus cuspidata cells under laminar shear stress. A total of 65 intracellular metabolites were identified and quantified, using gas chromatography coupled to time-of-flight mass spectrometry. Potential biomarkers were found by the principal component analysis as well as partial least squares combined with variable influence in the projection. Trehalose, sorbitol, ascorbate, sucrose, and gluconic acid were mainly responsible for the discrimination between shear stress induced cells and control cells. Further analysis by mapping measured metabolite concentrations onto the metabolic network revealed that shear stress imposed restrictions on primary metabolic pathways by inhibiting tricarboxylic acid cycle, glycolysis, and N metabolism. To adapt to the shear condition, cells responded by starting defensive programs. These defensive programs included coinduction of glycolysis and sucrose metabolism, accumulation of compatible solutes, and antioxidative strategy. A strategy of defense mechanisms at the level of metabolites for T. cuspidata cells when challenged with the shear stress was proposed. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source]

Evaluation of CE methods for global metabolic profiling of urine

ELECTROPHORESIS, Issue 14 2010
Rawi Ramautar
Abstract In this study, the usefulness of noncovalently coated capillaries with layers of charged polymers is investigated to obtain global electrophoretic profiles of urinary metabolites covering a broad range of different compound classes in a highly repeatable way. Capillaries were coated with a bilayer of polybrene (PB) and poly(vinyl sulfonate) (PVS), or with a triple layer of PB, dextran sulfate (DS) and PB. The bilayer and triple layer coatings were evaluated at acidic (pH 2.0) and alkaline (pH 9.0) separation conditions, thereby providing separation conditions for basic and acidic compounds. A representative metabolite mixture and spiked urine samples were used for the evaluation of the four CE methods. Migration time repeatability (RSD<2%) and plate numbers (N, 100,000,400,000) were similar for the test compounds in all CE methods, except for some multivalent ions that may exhibit adsorption to oppositely charged coatings. The analysis of cationic compounds with the PB-DS-PB CE method at low pH (i.e. after the EOF time) provided a larger separation window and number of separated peaks in urine compared to the analysis with the PB-PVS CE method at low pH (i.e. before the EOF time). Approximately, 600 molecular features were detected in rat urine by the PB-DS-PB CE-MS method whereas about 300 features were found with the PB-PVS CE-MS method. This difference can be attributed to reduced comigration of compounds with the PB-DS-PB CE-MS method and a related decrease of ion suppression. With regard to the analysis of anionic compounds by CE-MS, in general analyte responses were significantly lower than that for cationic compounds, most probably due to less efficient ionization and to ion suppression effects caused by the background electrolyte. Hence, further optimization is required for the sensitive CE-MS analysis of anionic compounds in body fluids. It is concluded that the selection of a CE method for profiling of cationic metabolites in urine depends on the purpose of the study. For high-throughput analyses, the PB-PVS CE-MS method is favored whereas the PB-DS-PB CE-MS method provides a more information-rich metabolic profile, but at the cost of prolonged analysis time. [source]

Amino acid profiling in plant cell cultures: An inter-laboratory comparison of CE-MS and GC-MS

ELECTROPHORESIS, Issue 9 2007
Brad J. Williams
Abstract A CE-MS method for metabolic profiling of amino acids was developed and used in an integrated functional genomics project to study the response of Medicago truncatula liquid suspension cell cultures to stress. This project required the analysis of more than 500 root cell culture extracts. The CE-MS method profiled 20 biologically important amino acids. The CE-MS method required no sample derivatization prior to injection and used minimal sample preparation. The method is described in terms of CE and MS operational parameters, reproducibility of migration times and response ratios, sample preparation, sample throughput, and reliability. This method was then compared with a previously published report that used GC-MS metabolic profiling for the same tissues. The data reveal a high level of similarity between the CE-MS and GC-MS amino acid profiling methods, thus supporting these as complementary technologies for metabolomics. We conclude that CE-MS is a valid alternative to GC-MS for targeted profiling of metabolites, such as amino acids, and possesses some significant advantages over GC-MS. [source]

Opportunities and constraints in the adaptation of technology for the diagnosis of bacterial plant diseases , experience from Tanzania,

EPPO BULLETIN, Issue 3-4 2000
R. Black
In order to improve diagnostic services and plant quarantine capabilities in Tanzania, the techniques of semi-selective media, the BACTID system, metabolic profiling (Biolog), indirect enzyme-linked immunosorbent assays (ELISA) and polymerase chain reaction (PCR) were assessed for suitability with the existing facilities for the diagnosis and detection of plant-pathogenic bacteria of vegetables. Field-collected samples as well as farmers' own and commercial germplasm were used in studies involving Ralstonia solanacearum, Clavibacter michiganensis subsp. michiganensis and Xanthomonas campestris pv. vesicatoria in Solanaceae and X. c. pv. campestris in Brassicaceae. Each of the techniques was used successfully with one or more of the target pathogens; each had advantages depending on the speed, sensitivity and specificity required, as well as the costs of carrying out the diagnosis. However, constraints emerged relating to the use and disposal of materials such as plastic Petri dishes and toxic substances. The more familiar underlying constraints of high cost and poor availability of consumables and erratic water and electricity supply continued to present problems. These problems will be discussed in relation to the development of an integrated and sustainable approach to the provision of routine diagnostic services. [source]

Metabolomics-based systematic prediction of yeast lifespan and its application for semi-rational screening of ageing-related mutants

AGING CELL, Issue 4 2010
Ryo Yoshida
Summary Metabolomics , the comprehensive analysis of metabolites , was recently used to classify yeast mutants with no overt phenotype using raw data as metabolic fingerprints or footprints. In this study, we demonstrate the estimation of a complicated phenotype, longevity, and semi-rational screening for relevant mutants using metabolic profiles as strain-specific fingerprints. The fingerprints used in our experiments are profiled data consisting of individually identified and quantified metabolites rather than raw spectrum data. We chose yeast replicative lifespan as a model phenotype. Several yeast mutants that affect lifespan were selected for analysis, and they were subjected to metabolic profiling using mass spectrometry. Fingerprinting based on the profiles revealed a correlation between lifespan and metabolic profile. Amino acids and nucleotide derivatives were the main contributors to this correlation. Furthermore, we established a multivariate model to predict lifespan from a metabolic profile. The model facilitated the identification of putative longevity mutants. This work represents a novel approach to evaluate and screen complicated and quantitative phenotype by means of metabolomics. [source]

HIGH-RESOLUTION MAGIC ANGLE SPINNING NMR ANALYSIS OF WHOLE CELLS OF CHAETOCEROS MUELLERI (BACILLARIOPHYCEAE) AND COMPARISON WITH 13C-NMR AND DISTORTIONLESS ENHANCEMENT BY POLARIZATION TRANSFER 13C-NMR ANALYSIS OF LIPOPHILIC EXTRACTS,

JOURNAL OF PHYCOLOGY, Issue 3 2004
Matilde S. Chauton
Lipid composition in extracted samples of Chaetoceros muelleri Lemmermann was studied with 13C-NMR and distortionless enhancement by polarization transfer (DEPT) 13C-NMR, resulting in well-resolved 13C-NMR spectra with characteristic resonance signals from carboxylic, olefinic, glyceryl, methylene, and methyl groups. The application of a DEPT pulse sequence aided in the assignment of methylene and methine groups. Resonance signals were compared with literature references, and signal assignment included important unsaturated fatty acids such as eicosapentaenoic and docosahexaenoic and also phospholipids and glycerols. Results from the extracted samples were used to assign resonance signals in a high-resolution magic angle spinning (HR MAS) DEPT 13C spectrum from whole cells of C. muelleri. The NMR analysis on whole cells yielded equally good information on fatty acids and also revealed signals from carbohydrates and amino acids. Broad resonance signals and peak overlapping can be a problem in whole cell analysis, but we found that application of HR MAS gave a well-resolved spectrum. The chemical shift of metabolites in an NMR spectrum depends on the actual environment of nuclei during analysis, and some differences could therefore be expected between extracted and whole cell samples. The shift differences were small, and assignment from analysis of lipophilic extract could be used to identify peaks in the whole cell spectrum. HR MAS 13C-NMR therefore offers a possibility for broad-range metabolic profiling directly on whole cells, simultaneously detecting metabolites that are otherwise not detected in the same analytical set up and avoiding tedious extraction procedures. [source]

Clinical trial: pilot study of metformin for the treatment of non-alcoholic steatohepatitis

ALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 2 2009
R. LOOMBA
Summary Background, Non-alcoholic steatohepatitis (NASH) is a form of progressive fatty liver disease that is strongly associated with insulin resistance, which suggests that insulin sensitizing agents such as metformin may be beneficial for NASH. Aim, To assess the effects of metformin on insulin sensitivity, body composition, serum alanine aminotransferase (ALT) levels and liver histology in patients with NASH. Methods, Patients underwent liver biopsy, metabolic profiling and imaging studies before and at the end 48 weeks of metformin (2000 mg/day) therapy. The primary endpoint was a three-point improvement in the histological NASH activity index. Results, Of 28 patients enrolled, 26 (13 females; average age 44 years) completed 48 weeks of treatment and underwent repeat metabolic studies, imaging and liver biopsy. Thirty per cent achieved a histological response. Most patients lost weight, the average being 6 kg. There was a marked association between weight loss and improvements in NASH activity index and ALT levels (both, P < 0.01). Insulin sensitivity also improved, but the degree of change did not correlate with histological improvement. Conclusion, Metformin leads to improvements in liver histology and ALT levels in 30% of patients with NASH, probably by its effects in causing weight loss. [source]

Development and validation of a gas chromatography/mass spectrometry method for the metabolic profiling of human colon tissue

RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 4 2009
Mainak Mal
In this study, a gas chromatography/mass spectrometry (GC/MS) method was developed and validated for the metabolic profiling of human colon tissue. Each colon tissue sample (20,mg) was ultra-sonicated with 1,mL of a mixture of chloroform/methanol/water in the ratio of 20:50:20 (v/v/v), followed by centrifugation, collection of supernatant, drying, removal of moisture using anhydrous toluene and finally derivatization using N -methyl- N -trifluoroacetamide (MSTFA) with 1% trimethylchlorosilane (TMCS). A volume of 1,µL of the derivatized mixture was injected into the GC/MS system. A total of 53 endogenous metabolites were separated and identified in the GC/MS chromatogram, all of which were selected to evaluate the sample stability and precision of the method. Of the identified endogenous metabolites 19 belonging to diverse chemical classes and covering a wide range of the GC retention times (Rt) were selected to investigate the quantitative linearity of the method. The developed GC/MS method demonstrated good reproducibility with intra- and inter-day precision within relative standard deviation (RSD) of ±15%. The metabolic profiles of the intact tissue were determined to be stable (100,±,15%) for up to 90 days at ,80°C. Satisfactory results were also obtained in the case of other stability-indicating studies such as freeze/thaw cycle stability, bench-top stability and autosampler stability. The developed method showed a good linear response for each of the 19 analytes tested (r2,>,0.99). Our GC/MS metabolic profiling method was successfully applied to discriminate biopsied colorectal cancer (CRC) tissue from their matched normal tissue obtained from six CRC patients using orthogonal partial least-squares discriminant analysis [two latent variables, R2Y,=,0.977 and Q2 (cumulative),=,0.877]. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Probing genetic algorithms for feature selection in comprehensive metabolic profiling approach

RAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 8 2008
Wei Zou
Six different clones of 1-year-old loblolly pine (Pinus taeda L.) seedlings grown under standardized conditions in a green house were used for sample preparation and further analysis. Three independent and complementary analytical techniques for metabolic profiling were applied in the present study: hydrophilic interaction chromatography (HILIC-LC/ESI-MS), reversed-phase liquid chromatography (RP-LC/ESI-MS), and gas chromatography all coupled to mass spectrometry (GC/TOF-MS). Unsupervised methods, such as principle component analysis (PCA) and clustering, and supervised methods, such as classification, were used for data mining. Genetic algorithms (GA), a multivariate approach, was probed for selection of the smallest subsets of potentially discriminative classifiers. From more than 2000 peaks found in total, small subsets were selected by GA as highly potential classifiers allowing discrimination among six investigated genotypes. Annotated GC/TOF-MS data allowed the generation of a small subset of identified metabolites. LC/ESI-MS data and small subsets require further annotation. The present study demonstrated that combination of comprehensive metabolic profiling and advanced data mining techniques provides a powerful metabolomic approach for biomarker discovery among small molecules. Utilizing GA for feature selection allowed the generation of small subsets of potent classifiers. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Reprogramming a maize plant: transcriptional and metabolic changes induced by the fungal biotroph Ustilago maydis

THE PLANT JOURNAL, Issue 2 2008
Gunther Doehlemann
Summary The fungal pathogen Ustilago maydis establishes a biotrophic relationship with its host plant maize (Zea mays). Hallmarks of the disease are large plant tumours in which fungal proliferation occurs. Previous studies suggested that classical defence pathways are not activated. Confocal microscopy, global expression profiling and metabolic profiling now shows that U. maydis is recognized early and triggers defence responses. Many of these early response genes are downregulated at later time points, whereas several genes associated with suppression of cell death are induced. The interplay between fungus and host involves changes in hormone signalling, induction of antioxidant and secondary metabolism, as well as the prevention of source leaf establishment. Our data provide novel insights into the complexity of a biotrophic interaction. [source]

Determination of urinary androgen glucuronides by capillary electrophoresis with electrospray tandem mass spectrometry

BIOMEDICAL CHROMATOGRAPHY, Issue 4 2009
Sung-Hee Cho
Abstract Capillary electrophoresis,electrospray tandem mass spectrometry (CE-ESI/MS/MS) is a simple and highly sensitive method for quantifying seven urinary androgen glucuronides. The urine samples were diluted and filtered through a membrane filter, and the filtrate was injected into a CE-MS/MS system without further sample preparation steps such as extraction and derivatization. The calibration ranges were 0.01,5 µg/mL for glucuronides of androsterone and 11, -OHAn-3G, and 5,500 ng/mL for glucuronides of 11-ketoAn, DHEA, testosterone, epitestosterone and DHT. The linearity of the method was 0.992,0.998, and the limits-of-detection at a signal-to-noise ratio of 3 were 5,10 ng/mL. The coefficients of variation were in the range of 4.0,9.0% for intra-day assay and 4.1,9.8% for inter-day assay. The proposed method may be applicable to metabolic profiling in both quantitative and qualitative analysis. Copyright © 2008 John Wiley & Sons, Ltd. [source]